Flashcards in Pathogenesis and Diagnostics Deck (75):
What are the three kinds of viral cytopathogenesis?
abortive, lytic, and persistent infections (chronic, latent, recurrent, and transforming)
What are the features of an abortive infection?
host is not permissive: temperature (viral replication better within certain ranges), inappropriate enzymes (lacks enzymes to uncoat, help w/ replication) or no receptor; or virus defective: missing genetic component for replication, need helper virus
What are the features of a viral lytic infection resulting in cell death?
morphologic changes: cytopathic effects (CPE)- membrane holes, expression or deletion membrane antigens, necrotic and degradative change in nuclear matter and protein, and inclusion bodies- aggregates of viral proteins, RNA or DNA, and altered host structures used diagnostically, cell fusion- syncytia or multi-nucleated giant cells
What virus is known for making multinucleated giant cells?
RSV respiratory syncytial virus
What viruses are known for their inclusion bodies?
rabies- Negri body, HSV- eosinophilic Cowdry type A body, CMV- basophilic nuclear inclusion body (owl inclusion)
What are the features of chronic non-lytic/persistent infection?
cell replicate many generations w/o releasing much virus, daughter cells contain virus from start, enough DNA or RNA produced to infect next cell, cells escape damage by patching, capping, and shedding decoy (surface proteins, immune system attacks decoy instead of cell)
What are the features of a latent non-lytic/persistent infection?
some viruses multiply profusely (total host not killed), lot of host DNA/RNA expressed, integration into host -genome w/ DNA and Retroviruses, hide and seek with immune system
What are the features of recurrent non-lytic persistent infections?
reactivation of latent infections: immune status of host compromised, replication increased, ability to escape host factors
What are the features of transformation non-lytic persistent infections?
occur when there are changes in normal growth; removal of brakes- don't stop multiplying, loss of contact inhibition, enhancement of accelerators or GFs
What are the host defenses against viral infections?
natural barriers (innate immune system), antigen specific immune responses (Ab esp. neutralizing, cell mediated CD8+, and interferons)
What is the difference between neutralizing and non-neutralizing Ab?
neutralizing- Ab to VAP1 or the viral associated protein that binds cellular receptor allowing virus to enter; non-neutralizing- Ab to a VAP that does not bind the receptor for entry,
What role do interferons play in viral infections?
decrease infection in neighboring cell, either: IFN-gamma TH1 responses by activating cell mediated immune response or IFN- alpha or beta to stop infection (unknown mechanism)
The course of viral infection is dependent on what factors?
nature of exporsure, immune status, age, general health, viral dose, and genetics of virus and host ( some people are susceptible no matter what and others are resistant no matter what)
what are the clinical phases of viral infection?
incubation period (time when virus gains access to cell and starts to produce virus), prodrome (non-specific symptoms prior to disease presentation, usually infectious), and disease (clinical illness)
What are some places or things that affect exposure?
vocation (day care, health workers, and teachers have greater exposure to different viruses), life-style (poor hygiene, drugs, alcoholism-destroy immune response), and living arrangements (children with elderly, infected spouses)
What are the potential routes of transmission?
direct contact, injection, transfusion, transplantation, fecal-oral (most common), and respiratory (i.e. sneeze)
What are some factors that transmission is dependent upon?
source, tissue site of replication and secretion, ability to endure hazards and barriers (environment and immune response), enveloped or not (major structural comp. determining transmission)
What is the difference between enveloped and non-enveloped viral survival?
non-enveloped- withstand drying, detergents, extreme pH and temperature; enveloped will not
What factors determine maintenance of virus in a population?
dependent on number of people not immune (VACCINATE!!) HERD immunity 70-90%
What factors effect the epidemiology of a virus?
exposure, transmission, maintenance of virus in population, age, geographic and seasonal considerations
What factor does age play in epidemiology of a virus?
infants and children generally more susceptible to all viruses, elderly more susceptible to new viruses or reactivations, mid-range- more severe disease with first exposure, great immune response causes most tissue damage and symptoms
What are the seasonal differences with viral infections?
Respiratory in fall and winter; Enteric- spring and summer
What is an outbreak?
a new virus in a new location
What is an epidemic?
over a large geographical area resulting from introduction of new strain of virus
What is a pandemic?
world-wide epidemic, occurs approximately every 5 years
What do most diagnostic virology tests demonstrate?
whether a specific virus is present or not; no test if there is any virus present (too many structural and genetic variations)
What is an indirect diagnostic viral test?
presumptive; test for immune consequences of virus infection
What is an direct diagnostic viral test?
conclusive; test for virus, viral protein or nucleic acid
Why are clinical signs and symptoms often used instead of diagnostic virology?
some presentations are definitive (Parvo-slapped cheek rash or varicella rash), during an outbreak (patients with same symptoms likely have same infection), some tests take too long or cost too much
What are the different direct methods of viral infection detetction?
culture-live virus, electron microscopy for virus structure, antigen, DNA/RNA genome
What are the features culturing a live virus?
cell culture, expensive and time consuming, not always possible, usually definitive, have to consider tropism (cell line expressing viral receptor allowing replication)
How is presence of virus in cell culture detected?
by cytopathic effect (CPE), varies by cell type and virus, requires expert interpretation; morphology of CPE helps but generally another test of culture is required to define viral type
what are the features of histology and cytology of patient specimens in viral detection?
look for characteristic changes in cell/tissue appearance, non-specific changes, require antibody stain for confirmation (skin, cervix, lavage, or lung washings), some viral inclusions can be diagnostic (HHV-6, BK, JC, adenovirus, or CMV)
What are the features of electron microscopy in viral detection?
require culture and concentration of virus or a patient sample with high levels of virus, allows ID of viral family but not serotype
What are the features of hemagglutination for viral detection?
used to titer (determine [viral]) viruses which bind RBC, serial dilutions added to RBC, sufficient virus= shield, low concentration = button; HA (hemagglutination units) least amount of virus required to create a shield.
What are the features of antigen detection in viral testing?
use virus specific antibody conjugated to enzyme (commercial prep) to detect virus proteins in tissue or in preparation (ELISA w/ indicator or by EIA- enzyme immune assay-
What is a shell vial?
detector cells grown in small vial (shell), inoculated with specimen, short term culture, confirmed by Ab, results 24-48 hours, easier than cell culture, like ELISA or EIA-Ab on membrane captures Ag in specimen, 2nd Ab carries enzyme that reduces substrate and produces color
What is direct fluorescence assay?
material collected from patient on glass slide, incubate with specific Ab to known virus (conjugated with fluoro) and washed, Ab binding detected by flourescene)
How is detection of viral genome by hybridization achieved?
use genomic material of virus as a target (RNA or DNA), following extraction from clinical sample (Southern, Northern slot or dot blots), relatively lower sensitivity, DNA/RNA dotted on membrane, electrophoresed on gel to separate sized, then detected by hybridization of tagged probe to nucleic acid
What are come features of PCR and RT-PCR with viral detection?
converts as little as one molecule of DNA to millions of copies, extremely sensitive and specific, PCR amplifies DNA, Reverse transcriptase-polymerase for RNA requires copying RNA as cDNA prior to amplification; amount DNA doubled with each cycle (quantitative)
What are the general features of indirect viral detection?
detect anti-viral Ab (patient sera), detect infection by virus w/o biopsy, can define serotype, use IgM (current infection) vs IgG (past infection or immunization, titers rise and fall w/ stage of infection), determine if reactivation
What is serology testing for?
capture Ab to viral antigens or related proteins, recombinant proteins or intact virus
How is serology tested?
ELISA/EIA (prep of viral antigens to capture anti-viral Ab), IFA (indirect immunofluorescence assay)
How is an IFA preformed?
virally infected cells fixed to glass slide, incubated with patient serum, antiviral Ab adhere viral antigens in infected cells, if bound pt. Ab detected w/ antihuman monoclonal Ab typically conjugated to fluorescing compound, visualized under fluorescence microscope (multiple dilutions to determine titer=  at which intensity staining diminishes)
What are differences in acute and convalescent titers?
1st titer drawn as soon as symptoms apparent, 2nd (convalescent) drawn 3-4wks later= traditionally 4 fold or greater increase in IgG; used to determine past infection or immunization and establish current resistance to infection
What are the features of an EBV (infectious mononucleosis) test?
heterophile Ab test for non-specific IgM produced early in disease (infected B cells); IgM Ab to VCA (viral capsid) specific for acute EBV; absence of anti-EBNA (nuclear) and other EBV Ab= recent infection; present anti-EBNA=infect. 3-4months prior
How is a western blot prefromed with viral indirect testing?
prepared viral proteins separated by gel electrophoresis, viral proteins transferred to membrane, Pt Ab detected by binding to protein
Give an example where indirect and direct viral tests in a panel are helpful.
HIV: blood bank preforns HIV NAT (direct nucleic acid test, RT-PCR) and HIV Ab assay; may detect anti-HIV (rapid EIA), then confirmed by HIV western blot, IFA, or RNA assay (RT-PCR), if indeterminant then HIV RNA qualitative test performed, if positive quantitative preformed
How are specimen's collected for diagnostic virology?
skin lesion scrapping, nasopharyngeal swab, naso-washing, lumbar puncture
What are the various forms of collection devices?
viral swab (different then bacterial), clot tube for blood (serology), Herpes- direct detection of herpes in skin lesions, sterile container (collect nasal wash or other body fluids)
How are samples transported?
mix specimen with transport medium in collection tube (except urine and CSF, would dilute specimen), refrigerate to protect from temperature extremes,
What comes in a herpes direct detection kit? What happens in the lab?
swab to collect specimen, slide to inoculate specimen and a fixative to preserve the specimen for transport and testing; stained with fluorescent-labeled antibody, virus infected exhibit apple-green fluorescence (either HSV1 or HSV2 will fluoresce but not both)
How is direct fluorescent staining?
detect herpes pathogens, need proper staining reagent, use for varicella zoster and HSV
How are results visualized with direct detection?
electron microscope- resolving power of 0.4-1nm, only available method to visualize virus particles, either scanning (scan specimen surface) or transmission (transmits thru thin layer of specimen on copper grid)
What are the features of direct detection histopathology?
virus produce a unique characteristic pattern when infecting tissues (allows for presumptive ID by direct detection) ex.- herpes esophagitis, CMV pneumonia, condylomatus dysplasia by HPV
How are nucleic acid probe?
sensitive and specific reagents, capable of detecting viral genomic DNA or RNA, use on clinical specimens or from culture isolates
when are nucleic acid probes used?
CMV, HSV, Enterovirus, norovirus, Hep C, HIV1 and 2
What are the classic nucleic acid probe test methods?
extraction, separation (ds into ss unless ss detected), addition of specific bio probe to detect virus, hybridization (specific DNA w/ complementary viral sequence), binding of detection complex to hybridized probe, and conversion of detection complex to a visualized reaction
What are the cell tissue culture types?
viral growth in vitro (tissue culture cells or cell lines), numerous types, selection dependent on specific virus, cell lines grown in tubes and flasks, visualized by standard light microscope or inverted microscope flask
what is a continuous cell line?
can be passed from tube to tube when changing medium to maintain viability
What is a primary cell line?
cells can only be passed from tube to tube a finite number of times before death
What is an inverted microscope?
used to visualize tissue culture cells grown in a flask, note the location of objective lenses
Why use specialized slanting racks?
holds tissue culture cells in tubes, ensures medium continuously covers the living cells, incubate at 37C
What is cytopathic effect or cytopathogenic effect? How is it observed?
virus manifests their presence in susceptible tissue culture cells by producing degenerative changes in the cells; either light or inverted microscope
what are the features of CPE of RSV?
appears within 3-4 days, forms large fused cells called syncticia- multinucleated giant cells
what are the features of CPE of HSV?
enlarged or ballooned cells, refractive cells lyse and eventually fall off the glass/plastic surface
What are the features of viral serology?
mainstay for hepatitis viruses, EBV, rubella, west nile, and HIV, use when recovery in culture is difficult or impossible, detection of IgM Ab in single specimen, can be diagnostic, paired sera (acute and convalescent) generally used (IgG)
What are the monospot test features?
detect heterophile IgM antibodies, diagnosis of infectious mononucleosis (EBV), uses immunological reaction of agglutination, detect non-specific antibody that is known to agglutinate horse RBCs, false positive common
what is the function of IgG serology?
detect for sero-conversion, test for single convalescent serum of IgG, determine the immune status to disease (CMV, rubella, VZ and HIV), visualized via ELISA or EIA to dilute to test titter
Which herpes viruses can be and how are they tested?
HSV 1 and 2, VZV, EBV, and CMV (can become latent), confirm diagnosis of HSV with direct FA exam of skin lesion
What are a few features of HIV and how is it diagnosed?
HIV 1 and 2 cause of AIDS, retrovirus, RNA to cDNA integrated into host DNA, serologic methods to detect specific Ab, typical serologic pattern of HIV Ag (early presence of HIV Ag- P24, GP41, 120/160), diagnosis requires screen and a confirmation, EIA screening, confirmation Western Blot on all reactive sera (need 2 of 3 major bands)
What are the features of HPV and testing?
warts in humans (hands, feet, genitals), can be cultured in vitro, Koilocytes in epithelial tissue suggest (colonoscopy biopsy) have dark small nucleus with clear cytoplasm appearance called condylomatous dysplasia, 16 & 18 high cervical cancer risk
What are the features of influenza virus and diagnostics?
influenza A and B,asociated with epidemic and pandemics, immunization, rapid and accurate diagnosis best method to control infection nasal washing best method, sensitivity and specificity from <50% to 95%, high false positives and negatives, tissue culture gold standard, serotype for epidemiologic purpose
What pathogens can be screened on influenza diagnostic kit?
adenovirus, influenza A and B, parainfluenza type 1, 2, and 3, and RSV